Rubisco accumulation factor 1 (Raf1) plays essential roles in mediating Rubisco assembly and carboxysome biogenesis

Significance Cyanobacteria are keystone organisms in global carbon fixation. Their great carbon-assimilation capability arises from a specialized virus-like protein organelle, the carboxysome, which comprises hundreds of proteins that form a shell to encapsulate the CO 2 -fixing enzymes Rubisco and carbonic anhydrase. How do these proteins self-assemble to construct the defined architecture? Here we explore the significance of one assembly factor, Raf1, in Rubisco assembly and carboxysome formation. We show that Raf1 mediates Rubisco assembly; without Raf1, carboxysome proteins are prone to form intermediate assemblies and small carboxysome-like structures rather than intact carboxysomes. Our results suggest a model of the Raf1-mediated biogenesis of carboxysomes and provide advanced knowledge of carboxysome assembly and function, informing synthetic engineering of functional CO 2 -fixing organelles for biotechnological applications.